牦牛肝蛋白BGP1抗肝癌功效学研究

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	牦牛肝蛋白BGP1抗肝癌功效学研究
	郭洁
学位类型	硕士
导师	李天才
	2014-05
学位授予单位	中国科学院研究生院
学位授予地点	北京
学位专业	中药学
关键词	牦牛肝蛋白制备抗癌活性
摘要	本文以牦牛肝为研究对象，采用磷酸缓冲溶液提取法从新鲜牦牛肝中提取牦牛肝蛋白并进行制备以及抗肝癌活性研究。利用pH7.2的磷酸缓冲溶液进行提取，经DEAE-52液相制备分离系统制备，通过建立H22荷瘤小鼠动物模型,测定肿瘤抑瘤率、体重、肿瘤体积、免疫指数和血清中白细胞介素-2（IL-2）及肿瘤坏死因子(TNF-α)水平，以及对肿瘤组织进行切片经HE染色观察以确定牦牛肝蛋白的抗肿瘤作用。取得的实验结果如下： 1. 从新鲜牦牛肝中提取粗蛋白平均提取率为23.66%，制备BGP1得率为5.60%。 2. 制备所得蛋白经SDS-PAGE进行分子量鉴定，其分子量在60 000-70 000Da之间，制备获得BGP1 30g。 3. 低、中、高剂量BGP1（100mg/kg/d、200mg/kg/d和300mg/kg/d）对小鼠H22肿瘤生长抑制率分别为50.15%、59.20%、73.35%，其抑制率随剂量的增加而上升，呈明显的剂量依赖性。 4. BGP1给药组与模型组对照，小鼠脾指数显著升高，胸腺指数显著降低；阳性对照（CTX）组胸腺指数、脾指数均显著下降；BGP1给药组H22小鼠血清中IL-2水平显著升高；低剂量BGP1组TNF-α水平明显升高，中高剂量变化不明显。 5. 肿瘤组织的病理切片结果：模型组细胞排列较密集，体积大小不一，有的呈侵袭性生长，有坏死；阳性对照组肿瘤呈弥漫性分布，肿瘤细胞细胞核固缩，出现大片坏死；BGP1低剂量组肿瘤细胞核固缩，变小，包浆少，坏死显著，BGP1中高剂量组出现片状坏死，细胞间质淋巴细胞浸润，数量明显增多。
其他摘要	Yak liver protein is studied in this paper. The phosphate buffer solution extraction method was used to extract crude protein from fresh yak liver. After extraction BGP1 preparation was done through DEAE-52 cellulose-based liquid chromatography separation and preparation system and then antitumor effect of BGP1 was studied. BGP1 was extracted from Yak (Bos grunniens) liver with phosphate buffer solution (pH7.2) and prepared by DEAE-52 cellulose-based liquid chromatography separation and preparation system.The model of transplanted H22 hepatocarcinoma bearing mice were used to investigate the antitumor efficiancy of BGP1 in vivo. Tumor inhibition rate, mice body weight change, tumor volume and immune organ index were tested. The concentration of IL-2 and TNF-a in serum of H22-bearing mice were also tested with ELISA kits. The results are as follows: 1. With the phosphate buffer solution extraction, the extraction rate was 23.66%. Through DEAE-52 cellulose-based liquid chromatography separation and preparation system, BGP1 yield was 5.60%. 2. By SDS-PAGE electrophoresis the molecular weight of BGP1 was determined, which was 60,000-70,000 Da. After preparation, about 30g BGP1 was obtained. 3. Compared with model control group, the tumor weight in BGP1 treated groups and CTX treated group shrank significantly with a tumor inhibition rate of 50.15% for 100 mg/kg BGP1 group(P<0.05), 59.20% for 200 mg/kg BGP1 group(P<0.05),73.35% for 300 mg/kg BGP1 group (P<0.001) and 75.10% for CTX group (P<0.001). 4. The spleen weight of mice in BGP1 groups were higher than those of model control group (P<0.05 or P<0.001), on the contrary, the thyrus weight of mice in BGP1 groups were lower than those of model control group (P<0.05). As expected, CTX dramatically decreased the immune organ index as compared to model control group (P<0.001 or P<0.05).The level of IL-2 were remarkably up-regulated in H22tumor-bearing mice after BGP1 administration (P<0.01) compared with the model control group. The level of TNF-α was significantly increased only after BGP1 low dosage administration (P<0.01), there is no difference between BGP1 (200,300 mg/kg/d) groups and model group. 5. Pathological observation of tumor tissue shows that, in the model group, tumor cells were arranged tightly and vary greatly in size with necrosis emerged. Cells in CTX group showed spots of necrosis, cells nucleus condensed; in 100mg/kg BGP1 group, significant necrosis of tumor cells emerged, in 200mg/kg and 300mg/kg BGP1 group, tumor cells have an intervening stroma with characteristic lymphoid infiltrates, while they were badly necrotic.
文献类型	学位论文
条目标识符	http://210.75.249.4/handle/363003/4021
专题	中国科学院西北高原生物研究所
推荐引用方式 GB/T 7714	郭洁. 牦牛肝蛋白BGP1抗肝癌功效学研究[D]. 北京. 中国科学院研究生院,2014.

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